This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. The CLC chloride-transport proteins orchestrate the movement of chloride necessary for proper neuronal, muscular, cardiovascular, and epithelial function. The structure of an E. coli CLC (ClC-ec1) in complex with a Fab fragment has been determined (pdb 1OTS). In work funded by the NIH (1R01GM070773-01A2), we have discovered an inhibitor of CLC-ec1, DIDS, which has an apparent affinity of ~30 uM. In this work, we propose to determine the structure of the ClC-ec1/DIDS complex. The results of these experiments will provide the first structure of a CLC inhibitor binding site. This structure will facilitate the use of DIDS as a tool to probe the chloride-transport mechanism in CLC-ec1, and could also prove useful for mapping the DIDS-binding site in the eukaryotic homologs that are inhibited by DIDS. In addition, the structure may aid in designing higher-affinity inhibitors, which are sorely lacking for the CLCs.